1Division of Pulmonary and Critical Care Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland, United States of America.

Abstract

BACKGROUND:

Respiratory dysfunction is a major contributor to morbidity and mortality in aged populations. The susceptibility to pulmonary insults is attributed to "low pulmonary reserve", ostensibly reflecting a combination of age-related musculoskeletal, immunologic and intrinsic pulmonary dysfunction.

METHODS/PRINCIPAL FINDINGS:

Using a murine model of the aging lung, senescent DBA/2 mice, we correlated a longitudinal survey of airspace size and injury measures with a transcriptome from the aging lung at 2, 4, 8, 12, 16 and 20 months of age. Morphometric analysis demonstrated a nonlinear pattern of airspace caliber enlargement with a critical transition occurring between 8 and 12 months of age marked by an initial increase in oxidative stress, cell death and elastase activation which is soon followed by inflammatory cell infiltration, immune complex deposition and the onset of airspace enlargement. The temporally correlative transcriptome showed exuberant induction of immunoglobulin genes coincident with airspace enlargement. Immunohistochemistry, ELISA analysis and flow cytometry demonstrated increased immunoglobulin deposition in the lung associated with a contemporaneous increase in activated B-cells expressing high levels of TLR4 (toll receptor 4) and CD86 and macrophages during midlife. These midlife changes culminate in progressive airspace enlargement during late life stages.

CONCLUSION/SIGNIFICANCE:

Our findings establish that a tissue-specific aging program is evident during a presenescent interval which involves early oxidative stress, cell death and elastase activation, followed by B lymphocyte and macrophage expansion/activation. This sequence heralds the progression to overt airspace enlargement in the aged lung. These signature events, during middle age, indicate that early stages of the aging immune system may have important correlates in the maintenance of tissue morphology. We further show that time-course analyses of aging models, when informed by structural surveys, can reveal nonintuitive signatures of organ-specific aging pathology.

A. Representative immunohistochemical staining for nitrotyrosine in mice at 2 months, 8 months, 12 months and 20 months of age. Robust staining is evident by 8 months of age. Site of staining is in alveolar epithelial cells (arrowheads). N = 4–6 mice per group. Original magnification, 20×. B. Quantitative immunohistochemistry of nitrotyrosine staining shows enhanced staining in the 8 month old lung which progresses through later time points. N = 6 mice per group. C. Quantitative immunohistochemistry of TUNEL staining of lungs from aging mice shows increased cell death at 8 months of age which persists through later time points. N = 6 mice per group. Reported data are mean values +/− SEM from at least five mice for each group. *p<0.05, **p<0.01.

Nine profiles generated by random selection for difference from evenly spaced profiles are depicted. The profile whose peak corresponded to the onset of airspace enlargement is enlarged on the left. Red shows the top 200 genes. Blue shows the remainder of genes within the cluster.

A. Representative histograms depicting lymphocyte subsets identified in lungs of mice at indicated ages. An increase in CD19+ cells occurs at 12 months of age compared with earlier time points (2 and 8 months). N = 4–6 mice per time point. B. Relative proportion of lymphocyte subsets quantified by flow cytometry in lung mononuclear cells isolated from mice at designated ages. N = 4–6 mice per time point. C. Quantitative immunohistochemistry of macrophage abundance in lungs of aging mice. An increase in macrophage infiltration occurs at 12 months of age compared with 8 months. Data are mean +/− SEM. N = 3–6 mice per time point. D. IL12/23 and IL17 ELISA analyses of lung lysates from mice at designated ages. A reduction of IL17 levels is evident in the lungs of mice 8 months of age and older compared with 2 month old mice.